Neuronal circuits are composed of a great diversity of cell types and it is likely that each cell type carries out a specialized function. (See P. Sterling, “The Synaptic Organization of the Brain,” edited by G. M. Shepherd, Oxford University Press, Oxford, 1990). Therefore, as a prerequisite to understanding the function of a circuit, it appears necessary to map synaptic connections among different types of neurons, or, as proposed in the Crick publication, to map all connections made onto a given cell. (See F H. Crick, Sci. Am. 241 (3), 219 (1979)).
Following the knowledge which describes the use of fluorescent membrane probes (see I. C. Farber and A. Grinvald, Science 222, 1025 (1983)), photostimulation of neurons using caged glutamate (see E. M. Callaway and L. C. Katz, Proc. Natl. Acad. Sci. U.S.A. 90, 7661 (1993)) has greatly advanced this research program, generating high-resolution input maps of neurons in brain slices. (See id.; M. B. Dalva and L. C. Katz, Science 265 (5169) 255 (1994); G. M. Shepherd, T. A. Pologruto, and K. Svoboda, Neuron 38 (2), 277 (2003); C. Boucsein, M. Nawrot, S. Rotter et al., Journal of neurophysiology 94 (4), 2948 (2005); R. Kotter, D. Schubert, J. Dyhrfjeld-Johnsen et al., J Biomed Opt 10 (1), 11003 (2005); H. U. Dodt, A. Schierloh, M. Eder et al., Neuroreport 14 (4), 623 (2003); and S. Shoham, D. H. O'Connor, D. V. Sarkisov et al., Nature methods 2 (11), 837 (2005)). In this method, glutamate uncaging is achieved by focusing ultraviolet light at a particular position in the slice, while simultaneously recording intracellular responses from a neuron at a different location. By moving the uncaging beam systematically across the slice, one can map the territories that generate excitatory or inhibitory responses in the recorded cell. While being useful, this method likely suffers from the problem that, due to the inherent scattering of light in living tissue and the large uncaging area generated by one-photon excitation, the stimulated area contains more than one neuron. Thus, one-photon photostimulation has not revealed synaptic connections between cells, but instead connections between a particular territory and a recorded neuron.
Accordingly, there may be a need to address at least some of the deficiencies described herein.